Jan Watteyne

The FMRFamide-Like Peptide Family in Nematodes

In the three decades since the FMRFamide peptide was isolated from the mollusk Macrocallista nimbosa, structurally similar peptides sharing a C-terminal RFamide motif have been identified across the animal kingdom. FMRFamide-like peptides (FLPs) represent the largest known family of neuropeptides in invertebrates. In the phylum Nematoda, at least 32 flp-genes are classified, making the FLP system of nematodes unusually complex. The diversity of the nematode FLP complement is most extensively mapped in Caenorhabditis elegans, where over 70 FLPs have been predicted. FLPs have shown to be expressed in the majority of the 302 C. elegans neurons including interneurons, sensory neurons, and motor neurons. The vast expression of FLPs is reflected in the broad functional repertoire of nematode FLP signaling, including neuroendocrine and neuromodulatory effects on locomotory activity, reproduction, feeding, and behavior. In contrast to the many identified nematode FLPs, only few peptides have been assigned a receptor and there is the need to clarify the pathway components and working mechanisms of the FLP signaling network. Here, we review the diversity, distribution, and functions of FLPs in nematodes.

Evolutionarily conserved TRH neuropeptide pathway regulates growth in Caenorhabditis elegans.

Significance The hypothalamic neuropeptide TRH (thyrotropin-releasing hormone) is one of the major endocrine factors that regulate vertebrate physiology. For decades the general assumption has been that TRH neuropeptides are not present in protostomes, at least not in ecdysozoans, despite the presence of TRH receptor orthologs in these phyla. Here we identify a TRH-related neuropeptide–receptor pathway in the nematode Caenorhabditis elegans. TRH-like neuropeptides activate the C. elegans TRH receptor ortholog in cell-culture cells. Using RNAi and CRISPR/Cas9 reverse genetics, we discovered that TRH-related signaling in the pharyngeal system promotes C. elegans growth. Our study provides evidence of a functional TRH neuropeptide–receptor pathway in invertebrates, suggesting that TRH signaling had evolved in a bilaterian ancestor more than 700 million years ago. In vertebrates thyrotropin-releasing hormone (TRH) is a highly conserved neuropeptide that exerts the hormonal control of thyroid-stimulating hormone (TSH) levels as well as neuromodulatory functions. However, a functional equivalent in protostomian animals remains unknown, although TRH receptors are conserved in proto- and deuterostomians. Here we identify a TRH-like neuropeptide precursor in Caenorhabditis elegans that belongs to a bilaterian family of TRH precursors. Using CRISPR/Cas9 and RNAi reverse genetics, we show that TRH-like neuropeptides, through the activation of their receptor TRHR-1, promote growth in C. elegans. TRH-like peptides from pharyngeal motor neurons are required for normal body size, and knockdown of their receptor in pharyngeal muscle cells reduces growth. Mutants deficient for TRH signaling have no defects in pharyngeal pumping or isthmus peristalsis rates, but their growth defect depends on the bacterial diet. In addition to the decrease in growth, trh-1 mutants have a reduced number of offspring. Our study suggests that TRH is an evolutionarily ancient neuropeptide, having its origin before the divergence of protostomes and deuterostomes, and may ancestrally have been involved in the control of postembryonic growth and reproduction.

Mass spectrometric evidence for neuropeptide-amidating enzymes in C. elegans

Neuropeptides constitute a vast and functionally diverse family of neurochemical signaling molecules and are widely involved in the regulation of various physiological processes. The nematode Caenorhabditis elegans is well-suited for the study of neuropeptide biochemistry and function, as neuropeptide biosynthesis enzymes are not essential for C. elegans viability. This permits the study of neuropeptide biosynthesis in mutants lacking certain neuropeptide-processing enzymes. Mass spectrometry has been used to study the effects of proprotein convertase and carboxypeptidase mutations on proteolytic processing of neuropeptide precursors and on the peptidome in C. elegans. However, the enzymes required for the last step in the production of many bioactive peptides, the carboxyl-terminal amidation reaction, have not been characterized in this manner. Here, we describe three genes that encode homologs of neuropeptide amidation enzymes in C. elegans and used tandem LC-MS to compare neuropeptides in WT animals with those in newly generated mutants for these putative amidation enzymes. We report that mutants lacking both a functional peptidylglycine α-hydroxylating monooxygenase and a peptidylglycine α-amidating monooxygenase had a severely altered neuropeptide profile and also a decreased number of offspring. Interestingly, single mutants of the amidation enzymes still expressed some fully processed amidated neuropeptides, indicating the existence of a redundant amidation mechanism in C. elegans. All MS data are available via ProteomeXchange with the identifier PXD008942. In summary, the key steps in neuropeptide processing in C. elegans seem to be executed by redundant enzymes, and loss of these enzymes severely affects brood size, supporting the need of amidated peptides for C. elegans reproduction.

Corrigendum: the FMRFamide-like peptide family in nematodes

[This corrects the article on p. 90 in vol. 5, PMID: 24982652.].